Rapid generation of splicing reporters with pSpliceExpress.
Almost all human protein-coding transcripts undergo pre-mRNA splicing and a majority of them is alternatively spliced. The most common technique used to analyze the regulation of an alternative exon is through reporter minigene constructs. However, their construction is time-consuming and is often complicated by the limited availability of appropriate restriction sites. Here, we report a fast and simple recombination-based method to generate splicing reporter genes, using a new vector, pSpliceExpress. The system allows generation of minigenes within one week. Minigenes generated with pSpliceExpress show the same regulation as displayed by conventionally cloned reporter constructs and provide an alternate avenue to study splice site selection in vivo.
Kishore
Shivendra
S
Department of Molecular and Cellular Biochemistry, B283 Biomedical Biological Sciences Research Building, 741 South Limestone, University of Kentucky, College of Medicine, Lexington, KY 40536-0509, USA.
Khanna
Amit
A
Stamm
Stefan
S
D001483:Base Sequence; D003001:Cloning, Molecular; D004262:DNA Restriction Enzymes / Q000737:chemistry; D005091:Exons; D017930:Genes, Reporter; D005821:Genetic Techniques; D006801:Humans; D008957:Models, Genetic; D008969:Molecular Sequence Data; D012322:RNA Precursors / Q000235:genetics; D012326:RNA Splicing; D012333:RNA, Messenger / Q000378:metabolism; D011995:Recombination, Genetic
D012322:RNA Precursors; D012333:RNA, Messenger; D004262:DNA Restriction Enzymes